The long-term goal is to understand the fundamental basis of complement signaling in the eye, and how mi- sregulation in this process leads to pathology, to ultimately aid in the development of therapeutic approach- es for devastating blinding diseases. Age-related macular degeneration (AMD) occurs in two forms, dry and wet. Dry AMD is characterized by drusen, RPE damage, and photoreceptor cell loss. In some patients, the dry form can transition to wet AMD. Wet AMD, presents itself with chroidal neovascularization (CNV), lea- kage of these new vessels, and rapid photoreceptor loss. Recent genetic evidence has implicated variations in the complement inhibitory protein factor H (CFH), as well as in the genes for complement factor B (CFB), C2 and C3, as potential risk factors for the disease. A common environmental stressor in AMD is oxidative stress. Three pathways activate the complement system: the classical (CP), alternative (AP), and mannose- binding lectin pathway (LP); all three converge on the same down-stream cascade. Experiments from our own laboratory as well as others, using the laser-damage model of CNV, have suggested that AP signaling is required for CNV development; whereas conflicting evidence has been published in the involvement of the other pathways. AP activity was found to control the generation of the proangiogenic factor vascular en- dothelial growth factor (VEGF), required for triggering new vessel growth. Finally, we have shown that AP activation is involved in oxidative stress-mediated RPE dysfunction characterized by VEGF and MMP re- lease from RPE monolayers. Oxidative stress was found to sensitize the RPE to complement attack by re- ducing the levels of membrane-bound endogenous complement inhibitors. For this proposal we will be guided by our overall hypothesis that pathologic activation of the AP has direct effects on the RPE, generat- ing a permissive cellular environment for AMD pathology. This hypothesis will be tested in three aims, both in vivo as well as in RPE cultures (primary human and mouse RPE cells). Using mice in which different pathways of the complement cascade are disrupted and complement-depleted serum, we will examine the relative roles of complement activation mechanisms and determine whether AP activation is required or suf- ficient. To identify the source of AP proteins (i.e., liver or eye), tissue-specific transgenic mice are analyzed. Next, we will test whether complement activation is specific for CNV, or whether AMD pathologies related to oxidative stress in the Sod1-/- mouse require a hyperactive complement cascade for them to develop. Expe- riments will be performed to test whether VEGF is involved in mechanisms of complement-mediated injury. And finally, the hypothesis will be put to test in vivo. We will use complement inhibitory strategies using tar- geted inhibitors that block the complement cascade at different levels to interfere with CNV. Testing inhibi- tors will not only establish their therapeutic value, but in addition, elucidating their mechanisms in CNV mice will investigate the roles and contributions of the different complement components in CNV.